Loss of Intrinsic Control in Autoimmune T Helper Cells with Signaling Variants

描述（由申请人提供）：B细胞、CD4 T细胞和髓样细胞在系统性红斑狼疮（SLE）等自身免疫性疾病的发展中起关键作用，并参与自身抗体和细胞因子产生、组织损伤和释放自身抗原激活的病理刺激网络。我们的建议是集中在CD4 T细胞和骨髓细胞在自身免疫性疾病。新技术增加了我们对SLE等疾病的遗传基础的理解，并有助于确定强易感位点，如MHC。最近的全基因组测序揭示了人类遗传变异的详细目录，每个人大约有12000个基因变异。我们对SLE或其他自身免疫性疾病中免疫细胞信号通路失调以及基因变异（非同义snp）如何支持异常免疫特征的认识就不那么可靠了。在这里，我们结合分子、细胞和体内方法来了解CD4 T细胞的自身免疫特征，这些T细胞如何与髓细胞在刺激环中纠缠，以及患者体内微妙的单等位基因、非同义snp如何导致强大的自身免疫表型。在初步结果中，我们提出了一种新的小鼠模型Rasgrp1Anaef， Ras激活子Rasgrp1的点突变导致自身免疫表型。对T细胞发育的深入表征表明，Rasgrp1Anaef T细胞发育有效，Rasgrp1Anaef小鼠淋巴细胞不减少。相反，CD44hiCXCR5+PD1+ CD4+ T细胞由于基础Rasgrp1Anaef- mTOR信号的增加而异常积聚，骨髓细胞亚群的数量增加，小鼠表现出抗核抗体和蛋白尿。Rasgrp1Anaef/WT杂合小鼠也表现出自身免疫病理，因此一个Rasgrp1WT等位基因不能补偿Rasgrp1Anaef。我们展示了最近发表的一种自身抑制性Rasgrp1二聚体的晶体结构，并证明Rasgrp1Anaef可以与Rasgrp1WT蛋白形成二聚体，这为在SLE、自身免疫性（1型）糖尿病和Graves病患者中报道的单等位基因Rasgrp1变异如何产生生物学效应提供了一种作用机制。在这里，我们将研究异常积累的Rasgrp1Anaef T细胞的免疫特性（Aim 1）， T-、B-和髓系细胞之间的密切相互作用，以及Rasgrp1Anaef髓系细胞在Rasgrp1Anaef自身免疫特征中的作用（Aim 2），以及这个Rasgrp1Anaef小鼠模型如何作为单等位基因、非同构RasGRP1 SNP等位基因的范例（Aim 3）。这些密切相关但独立的研究结果将增加我们对T细胞和骨髓细胞如何通过病理刺激网络驱动自身免疫的理解，以及RasGRP1中的单等位基因SNP事件如何导致自身免疫病理。

DESCRIPTION (provided by applicant): B cells, CD4 T cells, and myeloid cells play a critical role in the development of autoimmune diseases such as systemic lupus erythematosus (SLE) and can be engaged in a pathologic stimulatory network of autoantibody and cytokine production, tissue damage, and activation by released self-antigens. Our proposal is centered on CD4 T- and myeloid- cells in autoimmune diseases. New techniques have increased our understanding of the genetic basis of diseases like SLE and helped identify robust susceptibility loci, such as MHC. Recent whole genome sequencing has revealed a detailed catalog of the human genetic variation with a very high occurrence of ~12,000 gene variants per person. Much less robust is our knowledge of dysregulated signaling pathways in immune cells in SLE or other autoimmune diseases and how gene variants (non-synonymous SNPs) may underpin abnormal immune features. Here we combine molecular-, cellular-, and in vivo- approaches to understand the autoimmune features of CD4 T cells, how these T cells are entangled in a stimulatory loop with myeloid cells, and how subtle monoallelic, non-synonymous SNPs in patients can result in robust autoimmune phenotypes. In Preliminary Results, we present a novel mouse model, Rasgrp1Anaef, with a point mutation in the Ras activator Rasgrp1 that causes an autoimmune phenotype. Thorough characterization of T cell development demonstrates that Rasgrp1Anaef T cells develop effectively and Rasgrp1Anaef mice are not lymphopenic. Instead, CD44hiCXCR5+PD1+ CD4+ T cells aberrantly accumulate due to increased basal Rasgrp1Anaef- mTOR signaling, numbers of myeloid cell subsets increase, and mice show ANA (anti-nuclear antibodies) and proteinuria. Rasgrp1Anaef/WT heterozygous mice also demonstrate autoimmune pathology, thus one Rasgrp1WT allele cannot compensate for Rasgrp1Anaef. We present a recently published crystal structure of an autoinhibitory Rasgrp1 dimer and demonstrate that Rasgrp1Anaef can form a dimer with a Rasgrp1WT protein, presenting a mechanism of action for how monoallelic RasGRP1 variants, reported in patients with SLE, autoimmune (Type 1) diabetes, and Graves disease, can have a biological effect. Here we will investigate the immune character of the aberrantly accumulating Rasgrp1Anaef T cells (Aim 1), the intimate interaction between T-, B-, and myeloid cells and the role of Rasgrp1Anaef myeloid cells in the Rasgrp1Anaef autoimmune features (Aim 2), and how this Rasgrp1Anaef mouse model may serve as a paradigm for monoallelic, non-synonymous RasGRP1 SNP alleles (Aim 3). Results from the strongly related but independent aims will increase our understanding of how T cells and myeloid cells with seemingly innocent and subtle SNP variants can drive autoimmunity through a pathological stimulatory network, and how monoallelic genetic SNP events in RasGRP1 may cause autoimmune pathology.